The goal of this project is to enrich for the induction and isolation of murine monoclonal antibodies with affinity for native envelope spikes of SIV. These experiments will take advantage of the phenomenon of immune tolerance to "focus" the immune response of immunized mice on particular regions or structures of the native envelope spike of SIVmac239 and HIV-1. The first specific aim of this project is to create strains of mice that are immunologically tolerant to typical, non-neutralizing determinants through transgenic expression of monomeric envelope protein. Transgenic lines will be created expressing SIV envelope sequences under the control of an inducible promoter. In experiments comprising the second specific aim, transgenic animals that are immunologically tolerant to viral envelope proteins will be used to "focus" the antibody response on epitopes present in the inoculum but not in the transgenic protein. To do this, animals will be immunized with chemically inactivated virion particles, in order to present the envelope in its native configuration (it is well established that broadly neutralizing antibodies against HIV and SIV need to have affinity for the native complex). The third specific aim is to isolate neutralizing mAbs using whole virion binding and neutralization assays as the primary screens. Inactivated virion immunogens to be used are 1) virions bearing heterologous epitopes in the context of the endogenous sequence and 2) virions with envelope identical to the endogenously expressed envelope. In the first case, HIV epitopes are presented on native SIV envelope complexes; because the transgenic host will be tolerant to SIV epitopes, the antibody response will be "focused" on the heterologous, HIV determinant. In the second case, the hypothesis to be tested is that native envelope complexes contain conformationally induced epitopes that are not present in the monomeric envelope protein.